1-methyl-2-isopropyl-5-nitroimidazole and water soluble salts thereof

ABSTRACT

THE PREPARATION OF 1-METHYL-2-ISOPROPYL-5-NITROIMIDAZOLE BY NITRATION OF 2-SIOPROPYLIMIDAZOLE TO FORM THE 2ISOPROYL-4(OR 5)-NITROIMIDAZOLE INTERMEDIATE WHICH THEN IS METHYLATED IS DISCLOSED. THE WATER SOLUBLE SALTS THEREOF ARE PREPARED BY REACTING WITH THE APPROPRIATE ACID. THE PRODUCTS HAVE ANT-HISTOMONAS AND ANTITRICHOMONAS PROPERTIES. COMPOSITIONS CONTAINING THE ACTIVE INGREDIENT ARE ALSO DISCLOSED.

United States Patent U.S. Cl. 260-309 1 Claim ABSTRACT OF THE DISCLOSUREThe preparation of 1-methyl-2-isopropyl-5-nitroimidazole by nitration of2-isopropylimidazole to form the 2- isopropyl-4(or 5)nitroimidazoleintermediate WhlCh then is methylated is disclosed. The water solublesalts thereof are prepared by reacting with the appropriate acid. Theproducts have anti-histomonas and antitrichomonas properties.Compositions containing the active ingredient are also disclosed.

RELATED APPLICATIONS This application is a division of US. patentapplication Ser. No. 817,179 filed Apr. 17, 1969, now US. Pat. No.3,502,776, which in turn is a continuation-in-part of U.S. patentapplication Ser. No. 541,968 filed Apr. 12 1966, now abandoned, thebenefit of the dates of which are claimed.

DETAILED DESCRIPTION This invention relates to a novel imidazolederivative useful in the veterinary field, to a novel intermediate inthe preparation thereof and to compositions containing said novelimidazole derivative. More particularly, the invention relates to1-methyl-2-isopropyl-5-nitroimidazole or its pharmaceutically acceptablewater-soluble acid addition salts or mixtures thereof and to their usefor promoting growth of turkeys, increasing feed efficiency in turkeysand for preventing and treating turkey blackhead.

According to the present invention it has been found that1-methyl-2-isopropyl-S-nitroimidazole possesses an unexpectedly highactivity against turkey blackhead disease, increases feed efiiciency andpromotes growth of the birds when administered to them at lowconcentrations in the diet. Turkey blackhead or enterohepatitis is anextremely fatal disease occurring in poultry, especially in turkeys. Itis produced by the organism Histomonas meleagridis which is carried inthe egg of the cecal worm Heterakis Gallinae.

The novel 1-methyl-2-isopropyl-S-nitroimidazole, its pharmaceuticallyacceptable water-soluble acid addition salts or mixtures thereof of thisinvention have been found to effect a complete cure of Histomonasmeleagridis infected turkeys when administered to provide the activecompound at levels as low as 0.005 percent by Weight of the feed ration.Prophylaxis can be obtained with even lower doses. For the effectivecontrol of enterohepatitis in turkeys the active compound of thisinvention can be administered at levels of from about 0.001 percent toabout 0.050 percent by weight of the diet. The preferred concentrationwill, of course, depend to some extent upon the severity of theinfection. In most cases, however, a turkey ration containing about0.003 percent to about 0.025 percent by Weight is suitable forprevention with 0.00625 percent preferred and 0.0125 to 0.025 ispreferred for therapy. When utilized in drinking water in the form ofits water-soluble acid addition salts, the levels "ice of activecompound administered are one-half the amount utilized in feed.

For increased feed efficiency and growth promotion in turkeys the activecompound of this invention can be administered at levels of from about0.001 percent to about 0.05 percent. The preferred concentration will,of course, depend to some extent upon the condition of the turkeys.Usually for growth promotant purposes, the active compound is added tothe feed of the turkeys beginning with the first week of life. Generallyabout 0.003 percent to about 0.025 percent by weight is suitable withabout 0.00312 to about 0.00625 percent by weight preferred. Thus, if itis desired to promote growth only, the optimum effect is achieved with adosage of about 0.00312 percent in feed. If however, it is desired topromote growth and also combat turkey blackhead, the optimum effect isachieved with a dosage of about 0.00625 in feed.

In addition to its use against turkey blackhead, the novel product ofthis invention, i.e., 1-methyl-2-isopropyl-5- nitroimidazole, is alsouseful in combating protozoal infections caused by a variety oforganisms among which there can be named E. histolytz'ca, Trichomonasvaginalis and Trichomonas foetus.

The novel imidazole of this invention is prepared by nitrating2-isopropylimidazole to give 2-isopropyl-4-(or 5 )nitroimidazole. Thenitration is suitably carried out by adding a nitrating agent, e.g., amixture of nitric and sulfurc acids, to the 2-isopropylimidazolestarting material. The reaction is conveniently carried out attemperatures in the range of about 0 C. to 100 C. though higher or lowertemperatures could also be employed.

The 2-isopropyl-4(or 5 )nitroimidazole obtained by the above-describednitration procedure can be readily alkylated by the usual alkylationtechniques such as, for example, by treating with dimethylsulfate in aninert solvent preferably at an elevated temperature to form the desired1-methyl-2-isopropyl-S-nitroimidazole. The 2-isopropy1- 4(or5)nitroimidazole intermediate is also a novel compound and thusconstitutes a part of this invention.

The novel, pharmaceutically acceptable acid addition salts of1-methyl-2-isopropyl-S-nitroimidazole are formed by the usualtechniques, preferably by reacting with a strong mineral acid in organicmedia, e.g., isopropanol. Typical suitable mineral acids are, e.g.,sulfuric acid, hydrochloric acid, nitric acid. Preferred arehydrochloric acid which forms the water-soluble hydrochloride salt andsulfuric acid which forms the Water-soluble bisulfate salt.

The novel imidazole compound of this invention is normally administeredto the turkeys intimately mixed in the turkey ration or the drinkingwater. The drug can be suitably prepared as a premix or feed supplementcontaining from about 1 percent to about percent by weight of the premixformulation which can also contain carriers or diluents such as cornmeal, germ meal, lactose, corn starch, talc, gelation, magnesiumstearate, oyster shell flour, calcium silicate and the like. Othercompatible medicaments may also be added to the premix. It has beenfound that the preferred diluents and carriers are oyster shell flourand calcium silicate since they form stable premixes which have goodpotency retention. The premix can be added to commercial feed andintimately mixed therewith to obtain uniform distribution yielding aneffective concentration level of the active compound, l-methyl-2-isopropyl-5-nitroirnidazole, for preventive and therapeutic use aswell as for increasing feed efficiency and promoting growth ranging fromabout 0.001 percent to about 0.05 percent of the commercial feed. Thefeed supplement or premix containing the novel compounds of thisinvention can be readily mixed with the turkey ration by anyconventional technique for mixing feeds. For convenience in commercialuse, it has been found that premixes containing from about percent toabout percent by weight of the active compound are prefered with themost preferred, a premix containing about 12.5 percent by weight.

When administering the compound in drinking water, it has been foundconvenient to utilize water-soluble powders comprising the activecompound in the form of its water-soluble pharmaceutically acceptableacid addi tion salts, preferably the hydrochloride or bisulfate salts,The powder can be composed of only the water-soluble salt, or can be amixture of the salt of the active compound and water-soluble excipients,c.g., lactose, dextrose, tartaric acid. The powder can be added todrinking water to yield an eifective concentration level of the activecompound for combating turkey blackhead as well as pro moting growth inamounts ranging from 0.001 percent to 0.05 percent of the activecompound.

The following examples illustrate the invention. All temperatures are inC.

EXAMPLE 1 Preparation of 2-isopropyl-4.(or 5 )nitroimidazole2-isopropylimidazole, M.P. 130-135 (90 g.=0.816 mole) was added understirring and cooling with ice water to nitric acid, d.=1.4 (200 ml.) insmall portions at a temperature not exceeding Concentrated sulfuric acidwas dropped into the resulting solution under stirring and cooling withice water at 1020. The mixture was then heated gradually to atemperature of 80-85 at which the reaction became self-sustainingaccompanied by foaming and evolution of nitrous gases. The latter wereremoved with a current of nitrogen. The temperature was kept below 90 byoccasionally immersing the vessel in a prepared ice water bath. When(after 30 to 45 minutes) the gas evolution had ceased, the temperaturewas raised to 1l0-120 and the solution kept at that temperature for 30minutes. The solution was then poured into 300400 g. of ice andneutralized with ammonia to a pH of 4-5. After standing at 0 for 1-2hours, the crystallized product was filtered by suction, washed on thefilter twice with 100 m1. of cold water and dried at 90100. The2-isopropyl-4-(or 5)nitroimidazole thus obtained melted at 182183.

EXAMPLE 2 Preparation of 1-methyl-2-isopropyl-S-nitroimidazole2-isopropyl-4(or 5)nitroirnidazole (31 g.=0.2 mole), dioxane (70 g.) anddimethylsulfate (28 g.=0.22 mole) were heated on a steam bath underreflux for 45 minutes. The solvent was removed in vacuo on a steam bath,the residue dissolved in 20 ml. of water and the product precipitated bythe gradual addition of 80 g. of percent sodium hydroxide solution at 0.A small additional amount was obtained by extraction of the motherliquor with methylene chloride. The product melted at 60.

The product was purified as follows:

60 g. of product was dissolved in 3 N aqueous hydrochloric acid, thesolution was treated with charcoal and filtered. The filtrate wasneutralized by the gradual addition of aqueous concentrated ammonia at0.5 under stirring whereupon the product precipitated in white plates inthe measure the neutralization proceeded. The precipitate was filteredby suction, washed on the filter with 50 ml. of ice cold water and driedat room temperature, MP. 60.

The hydrochloride salt was formed by reacting the product, dissolved inisopropanol, with 25 percent ethanolic hydrochloric acid, whereupon thesalt precipitated and was isolated. It has a melting point of 177182dec. Similarly, the bisulfate salt was formed using 96 percent sulfuricacid. It has a melting point of 151.5- 152.5".

4 EXAMPLE 3 The following example illustrates typical feed supplementformulations embodying the novel 1-methyl-2-isopropyl-S-nitroimidazoleof this invention.

1 methyl 2 isopropyl-S-nitroimidazole 6 percent premix for animal useGrams/kilo 1-methyl-2-isopropyl-5-nitroimidazole 62.5

Pulverized oyster shell flour 887.5 Microcel E (calcium silicate) 50Total weight (gms.) 1000.0

Procedure (1) The pulverized oyster shell flour was placed in a suitablemixer and while mixing, the Microcel E was slowly added and thoroughlymixed.

.(2) While mixing continuously the 1-methyl-2-isopropyl-S-nitroimidazolewas slowly added and mixed until the mixture was homogeneous.

(3) This premix was then added to a commercial poultry feed at the ratioof 4 lbs./ton to yield a ratio of about .0125 percent drug andthoroughly mixed.

(4) This medicated feed was used in the mash form and it was alsopelleted on a Sprout-Waldron pellet mill.

Amounts of the above premix may be added to the commercial feed to yieldmedicated levels ranging from 0.001 percent to 0.05 percent. Thecommercial feeds to which this premix is added may be free of othermedicaments or may contain other medicaments if the final mixture iscompatible.

1-methyl-2-isopropyl-S-nitroimidazole 12 /2 percent premix for animaluse Grams/kilo 1-methyl-2-isopropyl-5-nitroimidazole Pulverized oystershell flour 825 Microcel E (calcium silicate) 50 Total weight (gms.)1000 Procedure 1) The oyster shell flour was placed in a suitable mixerand while mixing, the Microcel E was slowly added and thoroughly mixed.

(2) While mixing continuously, thel-methyl-Z-isopropyl-S-nitro-imidazole was slowly added and mixed untilthe mixture was homogeneous.

(3) This premix was then added to a commercial poultry feed at the ratioof 1 lb./ton to yield a ratio of 0.00625 percent drug and thoroughlymixed.

(4) This medicated feed was used in the mash form and it was alsopelleted on a Sprout-Waldron pellet mill.

Amounts of the above premix may be added to the commercial feed to yieldmedicated levels ranging from 0.001 percent to 0.05 percent. Thecommercial feeds to which this premix is added may be free of othermedicaments or may contain other medicaments if the final mixture iscompatible.

l-methyl-2-isopropyl-5-nitroimidazole hydrochloride water-soluble powderformulation Grams 1-methyl-2-isopropyl-S-nitroimidazole hydrochloride14.3 Tartaric acid 4.0 Lactose 1.7

Procedure (1) The 1-methyl-2-isopropyl-S-nitroimidazole hydrochloride,dextrose, lactose were blended in a suitable mixer to form a homogeneouspowder.

(2) 20 grams of the powder was then added to 50 1 EXAMPLE 4 Ten turkeypoults for each of medicated group, infected unmedicated controls (IUC)and uninfected unmedicated controls (UUC) were employed. Indicatedamounts of the drug were mixed with the turkey feed in a mechanicalmixer and given 72 hours in advance of the infection. The infectionconsisted in administering orally by pipette 200 embryonated eggs ofHeterakis gallinae containing Histomonas meleagridis. The turkeys werekept on the test drug feed mixture for 24 days and then sacrificed andautopsied and examined for hepatic and cecal lesions. The number ofsurvivors and the intensity of lesions appearing in the liver and cecaof each turkey was observed. The percent mortality was recorded for eachgroup. The relative growth rate of the treated turkeys and uninfectedcontrols was determined and is expressed as the weight gain (WG) inpercent. 100 percent represents the growth rate of the uninfectedunmedicated controls. The results of experiments in which infectedturkeys were fed various concentrations of1-methyl-2-isopropyl-S-nitroimidazole are set forth in the followingtable.

WG (per- Mortality cent) (percent) Percent of active Drug in feed:

EXAMPLE 5 Ten turkeys, one day old, were used per group and ten turkeysfor the unmedicated controls were employed. Indicated amounts of thedrug were mixed with the turkey feed in a mechanical mixer. The turkeyswere kept on the test drug feed for four weeks. The records of weightgain, feed consumption and feed efliciency were kept. The results areshown in the following table.

WG (per- Feed elfieent) ciency Percent of active drug in feed:

EXAMPLE 6 drug feed mixture for 21 days and then sacrificed andautopsied and examined for hepatic and cecal lesions. The number ofsurvivors and the intensity of lesions appearing in the liver and cecaof each turkey was observed. The percent mortality was recorded for eachgroup. The relative growth rate of the treated turkeys and uninfectedcontrol was determined and is expressed as the weight gain (WG) inpercent. percent represents the growth weight of the uninfectedunmedicated controls. The results are set forth in the following table.

Treatment days after infection W G (percent) Mortality (percent) EXAMPLE7 Ten turkey poults for each of medicated groups, infected unmedicatedcontrols (IUC) and uninfected unmedicated controls (UUC) were employed.Indicated amounts of the drug were dissolved in drinking water and givento the birds seven days after infection. The infection was administeredthe same as in Example 6. The turkeys were kept on the test drug for 14days and then sacrificed and autopsied and examined as in Example 6. Theresults are set forth in the following table.

WG Mortality (percent) (percent) What is claimed is:

1. A compound selected from the group consisting of1-methyl-2isopropyl-S-nitroimidazole and pharmaceutically acceptableWater-soluble acid addition salts thereof.

References Cited 23-9 (1966, January issue), RS1.

NATALIE TROUSOF, Primary Examiner U.S. Cl. XJR. 99-4; 424-273

